Ubiquitin-mediated proteolysis is an important pathway of non-lysosomal protein degradation which controls the destruction of many cellular regulatory proteins including p27, p53, p300, cyclins, E2F, STAT-1, c-Myc, c-Jun, EGF receptor, IkBa, NfkB and b-catenin (Pagano (1997) FASEB J. 11:1067). Ubiquitin is a highly conserved 76-amino acid polypeptide that is present in eukaryotic cells. The ubiquitin pathway leads to the covalent attachment of a polyubiquitin chain to target substrates that are then degraded by a multi-catalytic proteasome complex (Goldberg et al. (1995) Science 269:682–685). Many of the steps regulating protein ubiquitination are known. Initially, the ubiquitin activating enzyme (E1) catalyzes the formation of a high-energy thioester bond with ubiquitin, which is then transferred to a reactive cysteine residue of one of many ubiquitin conjugating enzymes. The final transfer of ubiquitin to an e-amino group or a reactive lysine residue in the target protein occurs in a reaction that need not require an ubiquitin ligase (E3) protein.
p27/Kip1 is a cyclin-dependent kinase (CDK) inhibitor that is predominantly regulated through the ubiquitin-mediated proteolytic pathway. The degradation of the regulatory protein p27/Kip1 appears to be required for G1-to-S phase transition (Sheaffet al. (1997) Genes Dev. 11:1464–1478). In both S-phase kinase-associated protein 2 (SKP2) and cyclin-dependent kinase subunit 1 (CKS1) knockout mice, p27/Kip1 was accumulated to high levels and proliferating cells were arrested in G1 to S-phase transition. Additionally, overexpression of p27/Kip1 in Hela cells resulted in growth inhibition that was associated with cell cycle G1 arrest (Tang and Nordin (1997) Bioch. and Biophys. Res. Comm. 238:534–538). Overexpression of p27/Kip1 also induced cell cycle arrest in G1 phase and subsequent apoptosis in HCC-9204 cell line (human hepatocellular carcinoma) and lung cancer (Yu et al. (1998) PNAS 95: 11324–11329). Furthermore, overexpression of p27/Kip1 has an anti-angiogenesis effect (Goukassian et al. (2001) FASEB J. 15:1877–1885).
The phosphorylation of p27/Kip1 on Thr187 by CDK2 creates a binding site for a SKP2 containing E3 ubiquitin-protein ligase known as skp1-cull-f-box (“SCF”) protein. Subsequent ubiquitination of p27/Kip1 by SCF results in the degradation of p27/Kip1 by the proteasome complex (Alessandrini et al. (1997) Leukemia 11:342–345). Additionally, SKP2, which functions as the receptor component of the SCF1 ubiquitin ligase complex, binds to p27/Kip in conjunction with CKS1 only when Thr187 of p27/Kip1 is phosphorylated. This critical binding and interaction appears to be necessary for the ubiquitination and degradation of p27/Kip1. Thus, the modulation of the ubiquitination of p27/Kip1 by E3 ubiquitin-protein ligase, which subsequently leads to degradation of p27/Kip1, provides an opportunity for the treatment and prevention of cancer, neoplastic and other proliferative diseases.
In addition, compounds with the general ability to suspend cells at a point in the cell-cycle without adversely affecting the long-term viability of the cell are useful as preservatives of a cell, blood, tissue or an organ in need of such preservation. As much as 60% of stored human blood and blood-products can be lost due to the limited “shelf-life”. The degradation in biological products such as whole cells is a result of catabolic processes at the cellular level and is inversely proportional to the storage temperature. A compound that can arrest cells in the G1 phase can increase the “shelf-life” of biological products or allow the biological products to be stored or transferred at elevated temperatures without an increase in the catabolic rate. Thus, there remains a need for compounds with the ability to preserve biological products.
2.1. Cancer and Neoplastic Disease
Cancer affects approximately 20 million adults and children worldwide, and this year, more than 9 million new cases will be diagnosed (International Agency for Research on Cancer). According to the American Cancer Society, about 563,100 Americans are expected to die of cancer this year, more than 1500 people a day. Since 1990, in the United States alone, nearly five million lives have been lost to cancer, and approximately 12 million new cases have been diagnosed.
Currently, cancer therapy involves surgery, chemotherapy and/or radiation treatment to eradicate neoplastic cells in a patient (see, for example, Stockdale, 1998, “Principles of Cancer Patient Management”, in Scientific American: Medicine, vol. 3, Rubenstein and Federman, eds., Chapter 12, Section IV). All of these approaches pose significant drawbacks for the patient. Surgery, for example, may be contraindicated due to the health of the patient or may be unacceptable to the patient. Additionally, surgery may not completely remove the neoplastic tissue. Radiation therapy is effective only when the irradiated neoplastic tissue exhibits a higher sensitivity to radiation than normal tissue, and radiation therapy can also often elicit serious side effects. (Id.) With respect to chemotherapy, there are a variety of chemotherapeutic agents available for treatment of neoplastic disease. However, despite the availability of a variety of chemotherapeutic agents, chemotherapy has many drawbacks (see, for example, Stockdale, 1998, “Principles Of Cancer Patient Management” in Scientific American Medicine, vol. 3, Rubenstein and Federman, eds., Ch. 12, sect. 10). Almost all chemotherapeutic agents are toxic, and chemotherapy causes significant, and often dangerous, side effects, including severe nausea, diarrhea, bone marrow depression, immunosuppression, etc. Additionally, many tumor cells are resistant or develop resistance to chemotherapeutic agents through multi-drug resistance.
Therefore, there is a significant need in the art for novel compounds and compositions, and methods that are useful for treating or preventing cancer or neoplastic disease with reduced or without the aforementioned side effects. Further, there is a need for cancer treatments that provide cancer-cell-specific therapies with increased specificity and decreased toxicity.
Citation or identification of any reference in Section 2 of this application is not an admission that such reference is available as prior art to the present invention.